Extraction method of human body secretory IgA, the human body secretory IgA, and O-glycosylation measurement method thereof

An extraction method and a secretory technology, which are applied in the preparation methods of peptides, chemical instruments and methods, and anti-animal/human immunoglobulins, etc., which can solve the difficulty of source, the complicated steps of serum separation, and the inability to carry out extensive medical research. And other issues

Inactive Publication Date: 2016-12-14
THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

This requires the extraction of IgA, but in the existing extraction methods, the extraction of IgA requires serum extraction, and the source is difficult. At the sam

Method used

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  • Extraction method of human body secretory IgA, the human body secretory IgA, and O-glycosylation measurement method thereof
  • Extraction method of human body secretory IgA, the human body secretory IgA, and O-glycosylation measurement method thereof
  • Extraction method of human body secretory IgA, the human body secretory IgA, and O-glycosylation measurement method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0108] Embodiment 1 collects human oral cavity secretion

[0109] The collected subjects rinsed their mouths with mineral water before brushing their teeth in the morning, and avoided drinking or eating for at least 30 minutes; gave the patients sterile cotton balls, chewed and secreted about 20ml of saliva, put them into sterile containers, and sent them to the experiment in a low-temperature insulation bucket. Centrifuge at a temperature of 4000r / min for 15min at 4°C, and collect the supernatant and store it in a -80°C refrigerator.

Embodiment 2

[0110] The preparation of embodiment 2 Jacalin-agarose affinity chromatography eluent

[0111] Eluent A: take NaCl 8.5g, NaCl 2 HPO 4 12H 2 O 2.9g, NaH 2 PO 4 .2H 2 O 0.3g was placed in a 2000ml beaker, 1000ml of deionized water was added, fully stirred and dissolved, and the eluent A solution was obtained.

[0112] Eluent B: Take 3.602g of melibiose and place it in a 200ml beaker, add 100ml of eluent A, stir well to dissolve, and obtain eluent B.

Embodiment 3

[0113] The packing column of embodiment 3 Jacalin-agarose affinity chromatography column

[0114] The packing method of Jacalin-agarose affinity chromatography column comprises the following steps:

[0115] Step (1): wash the column material, take 5ml, 2 times of immobilized lectin, wash with deionized water for 3 times, then wash with 0.1M PBS buffer solution once, and then soak the column material in 20% ethanol;

[0116] Step (2): Load the column material, first rinse the column body and the filter screen with ultra-clean water, slowly inject liquid A, then install the filter screen, fix the column material, and install the upper part of the column body after the column material sinks completely until it is close to Column material;

[0117] Step (3): Calculate the column volume, measure the height of the column material installed at last, and obtain a column volume of 8ml;

[0118] Step (4): Press the column, connect to the fast protein liquid chromatography, use the elu...

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Abstract

The invention belongs to the technical field of biology and provides an extraction method of human body secretory IgA, the human body secretory IgA, and an O-glycosylation measurement method thereof. The extraction method includes the steps of: firstly extracting oral cavity secretion of human body and centrifuging the oral cavity secretion; treating the oral cavity secretion to separate and purify the human body secretory immune globulin IgA by means of fast protein liquid chromatography with a Jacalin-agarose affinity chromatographic column; and performing the O-glycosylation measurement to the purified IgA. The method, compared with serum extraction, is painless, is convenient and is harmless to human body, and compared with urine IgA extraction, is high in content of the IgA after extraction and purification and is simple in separation and enrichment. The method can be widely applied in medical researching and laboratory researching.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for extracting human secretory IgA, a method for measuring human secretory IgA and its O-glycosylation. Background technique [0002] IgA protein, also known as immunoglobulin A, is second only to IgG in normal human serum, accounting for 10-20% of serum immunoglobulin content. According to immune function, it is divided into serotype and secretory type. Serotype IgA exists in serum, and its content accounts for about 85% of total IgA. Although serotype IgA has some functions of IgG and IgM, it does not show important immune functions in serum. Secretory IgA exists in secretions, such as saliva, tears, colostrum, nasal and bronchial secretions, gastrointestinal fluids, urine, sweat, etc. Secretory IgA is the main antibody in the local mucosal anti-infection immunity of the body. Therefore, it is also called mucosal local antibody. [0003] IgA nephropathy (IgA nephropat...

Claims

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Application Information

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IPC IPC(8): C07K16/18C07K1/34C07K1/22G01N33/68
CPCC07K16/18G01N33/6854
Inventor 张军军刘亚飞黄博余桂珍
Owner THE FIRST AFFILIATED HOSPITAL OF ZHENGZHOU UNIV
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